*3.4. Microfluidics*

A major challenge in gastrointestinal microbiome research is the existence of the microbiota as a community in which most members are ye<sup>t</sup> to be cultivated thus making it di fficult to identify what species is doing what within the ecosystem. Microfluidics technology is providing a platform where single microbial cells within the gastrointestinal microbiota can be tracked, studied and manipulated. Liu and Walther-Antonio [58] identified two powerful microfluidics that have the potential applications in cell sorting, cell culture, cell screening, genome application, metabolic screening/analyses and gene expression. A recent study by Chijiwa et al. [52] used single cell sequencing based on an SAG-gel platform that employed microfluidic droplet generator to unravel the metabolic function of uncultivated bacterial species in intestinal microbiome, in which the fermentation of dietary fiber resulting in the production of short-chain fatty acid from ingested fibers was evaluated. This technique enabled the thorough study of specific bacterial species and deciphering their specific function that contributes to the health of the gastrointestinal tract. Another interesting innovation in the area of microfluidics in microbiome research is the development of organs on chip. This has allowed the design of experiments that captures minute complexities of microenvironment with extraordinary resolution, giving details of the microbial diversity, structure and functions of the microbiota of specific organs of the human system [59]. The development of a primary human small intestine on a chip using biopsy derived organoids by Kasendra et al. [60] (as shown in Figure 2), in which they suggested that its potential use

in the study of infection, metabolism, drug pharmacokinetics and nutrition can be a viable and efficient tool in studying human gastrointestinal microbiota. Another principal application of microfluidics is in the study of antibiotic susceptibility of bacterial cells in real time. Cama et al. [61] in their study demonstrated the rapid quantification of antibiotic accumulation in Gram negative bacteria.

**Figure 2.** Primary human intestine chip manufacturing outline. (**a**) A cross-sectional view from the top of the chip and a phase contrast micrograph of the chip viewed from the bottom showing the upper (epithelial; blue) and lower (microvascular; pink) cell culture microchannels separated by a porous, Extracellular Matrix (ECM)-coated, Polydimethylsiloxane (PDMS) membrane sandwiched in-between. (**b**) An outline in developing microfuidic co-cultures of primary human intestinal epithelium and intestinal microvascular endothelium in the intestine chip. Source: Kasendra et al. [60] (*Scientific Reports*, Springer Nature) Licensed under CC BY 4.0.

By employing a combination of time-lapse auto-fluorescence microscopy and single-cell microfluidics, this technology could help reduce the overprescription of antibiotics, which is known to drive resistance. Beyond the aforementioned potential application of microfluidics, it also has clinical application in understanding the pathogenesis of gastrointestinal diseases, diagnosis, drug delivery and personalized or individualized medicine.
